Normal Rat Kidney Research Articles

A new flavonoid from the thorn of Gleditsia sinensis Lam

A new flavonoid, (2′′′E,6′′′S)-4′′-(6-hydroxy-2,6-dimethylocta-2,7-dienoyl)-vitexin (1), and five known compounds (2–6) were isolated from the thorn of Gleditsia sinensis Lam. Their structures were determined by comprehensive and comparative spectroscopic analysis of NMR and MS data. The absolute configuration of the new compound was deduced by analysis of the experimental and calculated 13C NMR data. The protective effects against lipopolysaccharide (LPS)-induced apoptosis in normal rat kidney tubule epithelioid (NRK 52e) cells of the isolated compounds were investigated in vitro, whose results showed that compound 1 showed significant protective effect with the EC50 value of 3.0 μM.

Molecular Changes in miRNA in Irradiated Rat Kidneys: Role of miR-34a and its Vascular Targets in the Notch Pathway.

The mechanism(s) of vascular regression in adult organs remains an unexplored gap. Irradiation to the kidney results in vascular regression and renal failure. The goal of this work was to determine molecular mechanism(s) of radiation-induced vascular regression and its mitigation by the drug lisinopril. Female WAG/RijCmcr rats received either 13 Gy X-ray irradiation, sparing one leg, or no irradiation, the latter serving as age-matched controls. Some irradiated animals received lisinopril. Kidney miRNA-seq was performed 35 days postirradiation, before symptoms of nephropathy. MicroRNA expression profiles were compared with data from humans. MicroRNA targets were predicted using TargetScan and confirmed by qRT-PCR and Western blot. Renal vascular endothelial cell density was evaluated at 100 days to confirm vascular regression. The normal rat kidney microRNA profile resembled that of humans. MiR-34a was increased >7-fold and emerged as the predominant rat microRNA altered by radiation. Expression of Jagged1, a ligand in the Notch pathway of vascular development and a target of miR-34a-5p was decreased by radiation but not in irradiated rats receiving lisinopril. Radiation decreased endothelial cells in the kidneys at 100 days, confirming vascular regression. In conclusion, the results of this study showed that radiation greatly increased miRNA34-a in rat kidneys, while lisinopril mitigated radiation-induced decrease of the Notch ligand, Jagged1, a molecular target of miRNA34-a.

Nociceptin Increases Antioxidant Expression in the Kidney, Liver and Brain of Diabetic Rats.

Simple SummaryNociceptin (NC) is a small peptide implicated in the physiology of pain, learning and memory. Here we investigated the role of NC in the induction of antioxidants in the kidney, liver, and the brain of diabetic rats using morphological and biochemical methods. Normal and diabetic animals were treated with NC for 5 days. Catalase (CAT) was expressed in the kidney, liver, and the neurons of the brain. Although CAT was markedly (p < 0.05) lower in the tubules of the kidney of normal and diabetic animals after NC treatment, NC significantly (p < 0.001) increased the presence of CAT in the liver and brain of diabetic rats. Superoxide dismutase (SOD) was observed in kidney tubules, hepatocytes, and neurons of the brain. Treatment with NC markedly (p < 0.001) increased the level of SOD in hepatocytes and neurons of the brain. Glutathione reductase (GRED) was seen in the convoluted tubules of the kidney, hepatocytes and neurons of the brain. Treatment with NC markedly increased (p < 0.001) the expression of GRED in kidney tubules, hepatocytes and neurons of the brain. In conclusion, NC can help diabetic patients mitigate the effects of oxidative stress by its ability to induce endogenous antioxidants.Nociceptin (NC) consists of 17 amino acids (aa) and takes part in the processing of learning and memory. The role of NC in the induction of endogenous antioxidants in still unclear. We examined the effect of NC on the expression of endogenous antioxidants in kidney, liver, cerebral cortex (CC), and hippocampus after the onset of diabetes mellitus, using enzyme-linked immunosorbent assay and immunohistochemistry. Exogenous NC (aa chain 1–17; 10 µg/kg body weight) was given intraperitoneally to normal and diabetic rats for 5 days. Our results showed that catalase (CAT) is present in the proximal (PCT) and distal (DCT) convoluted tubules of kidney, hepatocytes, and neurons of CC and hippocampus. The expression of CAT was significantly (p < 0.05) reduced in the kidney of normal and diabetic rats after treatment with NC. However, NC markedly (p < 0.001) increased the expression CAT in the liver and neurons of CC of diabetic rats. Superoxide dismutase (SOD) is widely distributed in the PCT and DCT of kidney, hepatocytes, and neurons of CC and hippocampus. NC significantly (p < 0.001) increased the expression of SOD in hepatocytes and neurons of CC and the hippocampus but not in the kidney. Glutathione reductase (GRED) was observed in kidney tubules, hepatocytes and neurons of the brain. NC markedly increased (p < 0.001) the expression of GRED in PCT and DCT cells of the kidney and hepatocytes of liver and neurons of CC. In conclusion, NC is a strong inducer of CAT, SOD, and GRED expression in the kidney, liver and brain of diabetic rats.

Oxidative Effects of Electromagnetic Radiation on Liver, Lung, Kidney and Heart Tissues of Diabetic and Normal Rats

Oxidative Effects of Electromagnetic Radiation on Liver, Lung, Kidney and Heart Tissues of Diabetic and Normal Rats

Renoprotective activity of a new amide and a new hydroxycinnamic acid derivative from the fresh roots of Rehmannia glutinosa

A new amide, named rehmagluamide (1), and a new hydroxycinnamic acid derivative, named nepetoidin F (2), together with six known compounds, 2′-O-methyluridine (3), puroglutamic acid (4), biliverdic acid (5), peterolactam (6), nicotinic acid (7), nicotinamide (8), were isolated from the fresh roots of Rehmannia glutinosa. All the structures of compounds were identified by the interpretation of their spectroscopic data and comparison with those reported in the literatures. The protective effects of compounds 1–7 on normal rat kidney tubule epithelioid (NRK-52e) cells injury induced by LPS were investigated. The results indicated that compounds 1, 2, and 7 exhibited protective effects against LPS-induced NRK 52e cells injury.

RAGE-MEDIATED CELLULAR SIGNALING IS TRIGGERED BY THE ALLOSTERIC ACTIVATION OF THE AT1 RECEPTOR

Objective: We recently identified the physical interaction of a pattern recognition receptor (PRR), lectin-like oxidized LDL receptor with the GPCR, angiotensin II type 1 receptor (AT1) whereby oxLDL allosterically activates AT1. In this study, we investigated if AT1 physically interacts with another PRR, the receptor for AGE (RAGE), and ligands of RAGE could allosterically activate AT1. Design and method: We used normal rat kidney epithelial cells (NRK52E) to investigate the interaction of the endogenous receptors. We also generated five different genetically-engineered CHO cells to individually analyze the G protein and β-arrestin pathways of AT1 activation; CHO cells expressing human RAGE (CHO-RAGE), CHO cells expressing both hRAGE and hAT1 (CHO-RAGE-AT1), CHO cells expressing both hRAGE and mutated hAT1 with impaired ability to activate G protein, and CHO cells expressing both hLOX-1 and mutated hAT1 with impaired ability to activate β-arrestin Results: In NRK52E, AGE-BSA provoked the activation of MAP kinase (ERK) that was inhibited by ARB or siRNA of AT1 or RAGE. Activation of NfκB and Epithelial mesanchmal transition induced by AGE-BSA was inhibited by ARBs as well as RAGE inhibitor. In genetically engineered CHO-cells, in situ PLA assay and immunoprecipitation confirmed the physical interaction of AT1 and RAGE on cellular membrane. HMGB1, a RAGE ligand induced Gαi-dependent inhibition of cAMP production only in the presence of both RAGE and AT1 with intact G protein binding. The effect of HMGB1 on cAMP content was abolished by ARB as well as RAGE inhibitor. HMGB1 did not induce the accumulation of IP1, a downstream of Gαq signaling, in CHO cells expressing both AT1 and RAGE. Real-time imaging of cellular membrane revealed that RAGE induced endocytosis of LOX-1 that depends on the intact β-arrestin pathway of AT1, and the phenomenon was not inhibited by ARB. Conclusions: RAGE ligands mediates cellular signaling via a physical interaction between RAGE and AT1. The activation of AT1 by RAGE ligands is distinct from that by angiotensin II in terms of selective G protein activation and the effect of ARB on G protein pathway but not on β-arrestin pathway.

Effects of Curcumin on High Glucose-Induced Epithelial-to-Mesenchymal Transition in Renal Tubular Epithelial Cells Through the TLR4-NF-κB Signaling Pathway.

ObjectiveDiabetic kidney disease (DKD) is a microvascular complication in diabetes mellitus, while tubuloepithelial to mesenchymal transition (EMT) of mature tubular epithelial cells is a key point in the early development and progression of renal interstitial fibrosis. The present study aimed to investigate the protective effects of Curcumin on EMT and fibrosis in cultured normal rat kidney tubular epithelial cell line (NRK-52E).MethodsBy using immunofluorescence staining and Western blot protocols, in vitro experiments were designed to analyze EMT markers, including collagen I and E-cadherin in high glucose (HG) exposed NRK-52E cells and to detect the expression levels of phosphorylated-NF-κB, TLR4 and reactive oxygen species (ROS) after Curcumin pre-treatment. With co-treatment with TAK242, these molecules in the TLR4-NF-κB signaling pathway were further evaluated.ResultsCurcumin decreased the HG-induced EMT levels and ROS production in NRK-52E cells. Furthermore, Curcumin was found to inhibit the TLR4-NF-κB signaling activation in HG-induced EMT of NRK-52E cells.ConclusionThe present study provides evidence suggesting a novel mechanism that Curcumin exerts the anti-fibrosis effects via inhibiting activation of the TLR4-NF-κB signal pathway and consequently protecting the HG-induced EMT in renal tubular epithelial cells. Thereby, TLR4-NF-κB may be a useful target for therapeutic intervention in DKD.

Resveratrol Attenuates Oxalate-Induced Renal Oxidative Injury and Calcium Oxalate Crystal Deposition by Regulating TFEB-Induced Autophagy Pathway.

The oxidative injury of renal tubular epithelial cells caused by inflammation and oxidative stress induced by hyperoxaluria is an important factor in the kidney calcium oxalate (CaOx) stone formation. Resveratrol (RSV) has been reported to reduce oxidative injury to renal tubular epithelial cells, and autophagy is critical for the protective effect of resveratrol. However, the protective mechanism of RSV in oxalate-induced oxidative injury of renal tubular cells and the role of autophagy in this process are still unclear. In our study, glyoxylic acid monohydrate-induced rats were treated with or without resveratrol, and it was detected that the overexpression of oxidant species, CaOx crystal deposition, apoptosis level, inflammatory cytokines and osteoblastic-associated protein expression were reversed by resveratrol. Additionally, Resveratrol pretreatment significantly reversed oxalate -induced decline in cell viability, cell damage, oxidant species overexpression, and osteogenic transformation in normal rat kidney epithelial-like (NRK-52E) cells. Furthermore, we found that RSV pretreatment promoted intracellular LC3II upregulation, p62 downregulation, and autophagosome formation, whereas 3-methyladenine treatment reduced this effect. Moreover, RSV induced the expression of transcription factor EB (TFEB) in the nucleus of NRK-52E cells in a concentration-dependent manner. After transfection of NRK-52E cells with TFEB siRNA, we showed that the RSV-induced increase in TFEB expression and autophagosome formation were inhibited. Simultaneously, RSV-induced NRK-52E cells protection was partially reversed. These results suggested that RSV regulates oxalate-induced renal inflammation, oxidative injury, and CaOx crystal deposition in vitro and in vivo through the activation of a TFEB-induced autophagy.

Ultrasonic Self-Emulsification Nanocarriers for Cellular Enhanced Astaxanthin Delivery.

Oil in water (O/W) nanocarriers were prepared for cellular enhanced astaxanthin delivery using a (3-carboxypropyl) triphenylphosphonium bromide (TPP)-modified casein by an ultrasonic self-emulsification method. The nanocarriers of casein emulsion loaded with astaxanthin and casein modified by TPP emulsion encapsulated with astaxanthin were 227 and 543 nm, respectively, with a spherical shape. The thermal stability and resistance to ultraviolet (UV) radiation ability of astaxanthin were significantly improved after encapsulation by the nanocarriers. The fluorescence colocalization imaging proved an accumulated effect of astaxanthin encapsulated in casein emulsion nanocarriers modified by TPP. Meanwhile, the astaxanthin loaded on TPP-modified nanocarriers could significantly protect the mitochondrial membrane potential from depolarization in the normal rat kidney (NRK) cells after oxidative damage. The cell viability assay demonstrated that the astaxanthin loaded on TPP-modified nanocarriers could enhance the growth of NRK and RAW264.7 cells as compared with astaxanthin encapsulated by casein emulsion without TPP modification.

3-Bromopyruvate decreased kidney fibrosis and fibroblast activation by suppressing aerobic glycolysis in unilateral ureteral obstruction mice model

AIMsEnhanced aerobic glycolysis is a motivation of fibroblast–myofibroblast transdifferentiation (FMT), leading to kidney fibrosis. 3-Bromopyruvate (3-BrPA) is a glycolysis inhibitor and has fibrosis-protected effect in liver. This study aims to explore the effects of 3-BrPA on aerobic glycolysis and kidney fibrosis in a unilateral ureteral obstruction (UUO) mice model and transforming growth factor-β1(TGF-β1)-stimulated normal rat kidney fibroblast (NRK49F) cell model in vitro. Main methodsIn vivo UUO mouse model and in vitro TGF-β1 stimulated cell model were built. Immunohistochemical staining, Western blots, Real-time PCR and fluorescence microscopy were employed to detect extra cellular matrix (ECM) synthesis, fibroblast activation, aerobic glycolysis switch and related signaling pathways. Key findingsHE and Masson's Trichrome staining showed that 3-BrPA substantially suppressed kidney injury and interstitial collagen production. 3-BrPA also attenuated ECM accumulation in a dose-dependent manner, as shown by immunohistochemistry staining, RT-PCR and western blot. Furthermore, 3-BrPA inhibited FMT, as indicated by α-SMA and PCNA immunofluorescence double staining. Additionally, the results of MTT assay indicated 3-BrPA prevented TGF-β1 induced fibroblasts proliferation in a time- and dose-dependent manner. Mechanistically, molecular docking results showed that 3-BrPA effectively decreased the aerobic glycolysis related enzymes Hexokinase-2 (HK-2), Lactate dehydrogenase A (LDHA) and Pyruvate kinase isozymes M2 (PKM-2), as well as inhibited IL-1 receptor–associated kinase 4 (IRAK4)/MYC protein levels. SignificanceOur study highlighted that 3-BrPA is a potential reno-protective agent in kidney fibrosis through the inhibition of fibroblasts aerobic glycolysis might via IRAK4/MYC signal pathways.

Non-cell-autonomous migration of RasV12-transformed cells towards the basal side of surrounding normal cells

Oncogenic transformation enables cells to behave differently from their neighboring normal cells. Both cancer and normal cells recognize each other, often promoting the extrusion of the former from the epithelial cell layer. Here, we show that RasV12-transformed normal rat kidney 52E (NRK-52E) cells are extruded towards the basal side of the surrounding normal cells, which is concomitant with enhanced motility. The active migration of the basally extruded RasV12 cells is observed when surrounded by normal cells, indicating a non-cell-autonomous mechanism. Furthermore, specific inhibitor treatment and knockdown experiments elucidate the roles of PI3K and myosin IIA in the basal extrusion of Ras cells. Our findings reveal a new aspect of cancer cell invasion mediated by functional interactions with surrounding non-transformed cells.

The Activation of ROS/NF-κB/MMP-9 Pathway Promotes Calcium-Induced Kidney Crystal Deposition.

Idiopathic hypercalciuria is an important risk factor for the formation of calcium-containing kidney stones. Matrix metalloproteinase-9 (MMP-9) is closely related to cell and tissue remodeling and is involved in ectopic tissue calcification. However, little is known about its role in kidney stone formation. In this study, we found that the expression of MMP-9 and that of osteoblastic-related proteins was increased in normal rat kidney epithelial-like (NRK-52E) cells following treatment with a high concentration of calcium, while the knockout or overexpression of MMP-9 could, respectively, significantly inhibit or upregulate the expression of osteoblastic-related proteins and calcium crystal deposition. In addition, apoptosis and calcium crystal deposition were significantly reduced in Sprague–Dawley rats with 1,25(OH)2D3-induced hypercalciuria following MMP-9 inhibitor I treatment. Furthermore, inhibiting reactive oxygen species (ROS) production or the nuclear factor kappa-light-chain-enhancer of activated B cell (NF-κB) pathway significantly reduced calcium-induced MMP-9 expression and calcium crystal deposition. In summary, our results suggested that a high calcium concentration promotes epithelial–osteoblastic transformation and calcium crystal deposition in renal tubule cells by regulating the ROS/NF-κB/MMP-9 axis and identified a novel role for MMP-9 in regulating calcium-induced calcium crystal deposition in renal tubules.

Nucleoporin 160 (NUP160) inhibition alleviates diabetic nephropathy by activating autophagy

ABSTRACT Diabetic nephropathy (DN) is the leading cause of end-stage renal disease worldwide. Autophagy was reported to be related to the pathogenesis of DN. This research investigated the function of the Nucleoporin 160 (Nup160) gene in regulating autophagy in DN. A mouse model of DN was established through an intraperitoneal injection of streptozotocin (STZ). Normal rat kidney tubular epithelial cells (NRK-52E) were treated with high glucose to induce DN in vitro. Real-time quantitative polymerase chain reaction (RT-qPCR), western blot, immunofluorescence assays were conducted to measure the expression of NUP160, autophagy-associated proteins, and inflammatory cytokines in vitro and in vivo. Pathological changes of kidney and liver tissues were analyzed using hematoxylin and eosin (H&E), Masson and periodic acid-silver (PAS) staining. The body weight, blood glucose, renal and lipid profiles of DN mice were examined. In this study, DN mice showed serious pathological injury. NUP160 expression was upregulated, autophagy was inhibited, and inflammatory response was increased in DN mice. Depletion of NUP160 restored autophagy and inhibited inflammation and fibrosis in high glucose (HG)-treated NRK-52E cells and STZ-induced DN mice by downregulating the expression of p62 and Collagen IV (Col-Ⅳ), increasing the ratio of LC3II/LC3I, and inactivating nuclear factor (NF)-κB signaling. Moreover, NUP160 knockdown could ameliorate pathological damage and glucose tolerance in DN mice. Overall, this study is the first to demonstrate the key role of NUP160 silencing in promoting autophagy against diabetic injury in DN.

Fluorescence nanoparticles from instant coffee accumulated in lysosome and induced lysosome-dependent cell death via necroptosis-like pathway

Fluorescence nanoparticles (FNs) are a type of nano-dots generated during baking process, and their safety on organism is unclear and little is known to their cytotoxicity. In this study, the FNs from instant coffee were purified and characterized. The FNs with an average size about 2.08 nm emitted bright blue fluorescence with lifetime about 2.74 ns. The element and functional groups were analyzed by X-ray photoelectron spectroscopy and Fourier transform infrared spectroscopy, respectively. The results indicated that these FNs were internalized in lysosomes and induced apoptosis of normal rat kidney (NRK) and Caco-2 cells. While, the pan-caspase inhibitor, Z-VAD-FMK didn't decrease the rate of apoptosis and cell death of the FNs-treated NRK and Caco-2 cells. These internalized FNs enlarged lysosomes, decreased lysosomal enzyme degradation activity and increased lysosomal pH value. Partial co-localization of receptor-interacting serine-threonine kinase 3 (RIPK3) to lysosomes in FNs-treated cells was observed, and the amount of RIPK1 and RIPK3 increased after treatment with FNs. The results demonstrated that the FNs from instant coffee induced lysosomal membrane permeabilization and initiated necroptosis.

Two new eremophilane-type sesquiterpenes from the fresh roots of Rehmannia glutinosa

• Two new eremophilane-type sesquiterpenes were isolated from the fresh roots of Rehmannia glutinosa . • The structures were established based on spectroscopic data analyses. • Compounds 1 and 2 exhibited protective effects against LPS-induced damage to NRK-52e cells. Two new eremophilane-type sesquiterpenes, remophilanetriol A ( 1 ) and remophilanetriol B ( 2 ), together with two known compounds, diincarvilone C ( 3 ) and diincarvilone D ( 4 ), were isolated from the fresh roots of Rehmannia glutinosa . The structures were determined based on analysis of the IR, UV, MS, and NMR data. The absolute configurations of the new compounds were resolved by comparing their experimental and calculated ECD spectra. All compounds were tested for their protective effects on normal rat kidney tubule epithelioid (NRK-52e) cells injury induced by lipopolysaccharide (LPS). The results showed that compounds 1 and 2 exhibited protective effects against LPS-induced damage to NRK-52e cells.

Contribution of TFEB-mediated autophagy to tubulointerstitial fibrosis in mice with adenine-induced chronic kidney disease

Autophagy has been implicated in the pathogenesis of chronic kidney disease (CKD). Transcription factor EB (TFEB) is a master controller of autophagy. However, the pathophysiological roles of TFEB in modulating autophagy and tubulointerstitial injury in CKD are unknown. This study aimed to determine whether TFEB-mediated autophagy contributed to the tubulointerstitial injury in mice with CKD. After the mice were treated with an adenine diet (0.2 % adenine) for 8 weeks, the development of CKD was observed to be characterised by increased levels of plasma blood urea nitrogen (BUN), creatinine (Cre), tubulointerstitial inflammation and fibrosis. Immunohistochemical and Western blot analysis further revealed that TFEB and autophagy genes were significantly up-regulated in the kidney of the mice with adenine-induced CKD, and this increase was mostly found in the tubular epithelial cells. Interestingly, a similar expression pattern of TFEB-autophagy genes was observed in tubular epithelial cells in the kidney tissue of patients with immunoglobulin A (IgA) nephropathy. Moreover, a pathogenic role of TFEB in adenine-induced CKD was speculated because the pharmacological activation of TFEB by trehalose failed to protect mice from tubulointerstitial injuries. In the epithelioid clone of normal rat kidney cells (NRK-52E), the activation of TFEB by trehalose increased autophagy induction, cell death and inflammatory cytokine (Interleukin-6, IL-6) release. Collectively, these results suggested that the activation of TFEB-mediated autophagy might cause autophagic cell death and inflammation in tubular epithelial cells, contributing to renal fibrosis in adenine-induced CKD. This study provided novel insights into the pathogenic role of TFEB in CKD associated with a high purine diet.

In Vitro Evaluation of the Apoptotic, Autophagic, and Necrotic Molecular Pathways of Fluoride.

Prolonged exposure to high doses of fluoride causes chronic poisoning called fluorosis, which affects many tissues and causes serious health problems. This study was planned to investigate the apoptotic, autophagic, and necrotic molecular pathways of fluoride. Sodium fluoride (NaF) was administered to normal rat kidney epithelial (NRK-52E) cells. The NaF IC50 value was determined using the MTT assay. The expression of the genes in the autophagic, apoptotic, and necrotic pathways was determined by real-time PCR. It was determined that there were significant changes in NaF-induced molecular pathways depending on the time. There were no increases in apoptotic and necrotic pathway markers except for Atg3, an autophagy gene, at the 3rd and the 12th hours. However, there was an induction in all cell death signaling pathways at 24 h. The molecular mechanisms demonstrated NaF-induced cellular death in the NRK-52E cell line. It was concluded that these molecular mechanisms were activated with NaF, and different mechanisms accelerated the cellular death at the 24th hour.

Proteomic landscape of TGF-\u03b21-induced fibrogenesis in renal fibroblasts

Transforming growth factor-β1 (TGF-β1) plays a premier role in fibrosis. To understand the molecular events underpinning TGF-β1-induced fibrogenesis, we examined the proteomic profiling of a TGF-β1-induced in vitro model of fibrosis in NRK-49F normal rat kidney fibroblasts. Mass spectrometric analysis indicated that 628 cell-lysate proteins enriched in 44 cellular component clusters, 24 biological processes and 27 molecular functions were regulated by TGF-β1. Cell-lysate proteins regulated by TGF-β1 were characterised by increased ribosomal proteins and dysregulated proteins involved in multiple metabolic pathways, including reduced Aldh3a1 and induced Enpp1 and Impdh2, which were validated by enzyme-linked immunosorbent assays (ELISA). In conditioned media, 62 proteins enriched in 20 cellular component clusters, 40 biological processes and 7 molecular functions were regulated by TGF-β1. Secretomic analysis and ELISA uncovered dysregulated collagen degradation regulators (induced PAI-1 and reduced Mmp3), collagen crosslinker (induced Plod2), signalling molecules (induced Ccn1, Ccn2 and Tsku, and reduced Ccn3) and chemokines (induced Ccl2 and Ccl7) in the TGF-β1 group. We conclude that TGF-β1-induced fibrogenesis in renal fibroblasts is an intracellular metabolic disorder and is inherently coupled with inflammation mediated by chemokines. Proteomic profiling established in this project may guide development of novel anti-fibrotic therapies in a network pharmacology approach.

SLC30A7 has anti-oxidant stress effects in high glucose-induced apoptosis via the NFE2L2/HMOX1 signal transduction pathway

AimsApoptosis and oxidant stress are known to be involved in the pathogenesis of diabetic kidney disease (DKD). We have previously reported that zinc transporter 7 in SLC30 family (SLC30A7) inhibits apoptosis in rat peritoneal mesothelial cells under high glucose (HG) conditions. In the current study, we aimed to investigate whether SLC30A7 had effect for anti-oxidant stress in renal tubular epithelial cells under HG. MethodsSLC30A7 in HG-induced apoptosis in a normal rat kidney tubular epithelial cell line (NRK-52E cells)/kidneys of STZ-induced diabetic mice was examined and the activity of nuclear factor erythroid 2-related factor 2 (NFE2L2) was further analyzed by using real time RT-PCR, siRNA and Western blot protocols. ResultsSLC30A7 was found to be up-regulated, while NFE2L2 was activated in kidneys of STZ-induced diabetic mice and HG-induced apoptosis of NRK-52E cells. Knock-down of SLC30A7 with siRNA protocol resulted in lower intracellular free zinc levels in the cells and decreased zinc distribution in the Golgi apparatus. Furthermore, knock-down of NFE2L2 down-regulated its target HMOX1 gene expression, decreased SLC30A7 activity but increased HG-induced apoptosis. ConclusionThe current study provides new evidence that SLC30A7 has anti-oxidant stress effects in HG-induced apoptosis via the NFE2L2/HMOX1 signal transduction pathway.

Synthesis and biological evaluation of substituted N-(2-(1H-benzo[d]imidazol-2-yl)phenyl)cinnamides as tubulin polymerization inhibitors

A new series of N-(2-(1H-benzo[d]imidazol-2-yl)phenyl) cinnamides was prepared and evaluated for their in vitro cytotoxic activity using various cancer cell lines viz. A549 (human non-small cell lung cancer), MDA-MB-231 (human triple negative breast cancer), B16-F10 (mouse melanoma), BT-474 (human breast cancer), and 4 T1 (mouse triple negative breast cancer). In the series of tested compounds, 12h showed potent cytotoxic activity against non-small cell lung cancer cell line with IC50 value of 0.29 ± 0.02 µM. The cytoxicity of most potent compound 12h was also tested on NRK-52E (normal rat kidney epithelial cell line) and showed less cytotoxicity compared to cancer cells. Tubulin polymerization assay indicated that the compound 12h was able to impede the cell division by inhibiting tubulin polymerization. Moreover, molecular docking study also suggested the binding of 12h at the colchicine-binding site of the tubulin protein. Cell cycle analysis revealed that the compound 12h arrests G2/M phase. In addition, 12h induced apoptosis in A549 cell lines was evaluated by various staining studies like acridine orange, DAPI, analysis of mitochondrial membrane potential, annexin V-FITC, and DCFDA assays.